Most antiviral drugs used to control the spread of HIV-1 have proven to be of limited use because under the selection pressure of the drug, the virus soon mutates to a drug-resistant strain. This tendency to develop drug resistance is a survival strategy used by many classes of viruses and is particularly pronounced among the members of the retrovirus family. One way to defeat this survival strategy is to focus on drugs attacking specific elements of the virus that are intolerant to mutations. Such elements can be identified by searching the proteins present in all viruses within the virus class to identify common or highly conserved structures.
A search of all known retroviruses reveals a highly conserved structure in their nucleocapsid (NC) proteins. All NC proteins of the Oncovirinae and Lentivirinae subfamilies of Retroviridae contain sequences of 14 amino acids with 4 invariant residues, Cys(X)2Cys(X)4His(X)4Cys (L. E. Henderson et al. J. Biol. Chem. 256, 8400 (1981)), which chelate zinc through histidine imidazole and cysteine thiolates with a Kd less than 10−3 (J. M. Berg, Science 232, 485 (1986); J. W. Bess, Jr., et al., J. Virol. 66, 840 (1992); M. R. Chance et al., Proc. Natl. Acad. Sci. U.S.A. 89, 10041 (1992); T. L. South and M. F. Summers, Adv. Inorg. Biochem. 8, 199 (1990); T. L. South, et al., Biochem. Pharmacol. 40, 123 (1990)) (see, FIG. 1). These structures are referred to as retroviral CCHC zinc fingers, and are one of the most highly conserved features of retroviruses. Examples of retroviruses which possess at least one CCHC type zinc finger per nucleocapsid protein include, but are not limited to, HIV-1, HUV-2, SIV, BIV, EIAV, Visna, CAEV, HTLV-1, BLV, MPMV, MMTV, RSV, MuLV, FeLV, BAEV, and SSV. The distribution of CCHC zinc fingers among known subgroups of retroviruses is indicated by the diagram set forth in FIG. 1. Due to their highly conserved nature, it is thought that CCHC zinc fingers perform an essential function in viral infectivity. In fact, it has been discovered that mutations of the chelating residues (CCHC) in the zinc fingers yield a non-infectious virus.
HIV-1 NC contains two zinc fingers separated by only 7 amino acids (L. E. Henderson et al., J. Virol. 66, 1856 (1992)). The location of the CCHC zinc fingers in the HIV-1 NC protein is indicated in the diagram shown in FIG. 2 (see also, Henderson et al., J. Virology, supra, 1992). The zinc fingers are not only required for packaging genomic RNA, but are also essential for early events in virus infection. Both fingers are essential for infectivity (A. Aldovini and R. A. Young, J. Virol. 64, 1920 (1990); R. J. Gorelick et al., J. Virol. 64, 3207 (1990).
Incubation of HIV-1 with 3-nitrosobenzamide (NOBA), a C-nitroso compound, results in the loss of zinc from the HIV-1 CCHC zinc finger array and viral inactivation (W. G. Rice et al., Nature 361, 473 (1993); W. G. Rice et al. PNAS 90, 9721 (1993)). Moreover, removal of zinc from the eukaryotic CCHC zinc finger of poly(ADP-ribose) polymerase by C-nitroso compounds has been reported (K. G. Buki, et al., FEBS Letters 290, 181 (1991)). Unfortunately, to date, the mechanism by which such CCHC zinc fingers are disrupted has not been discovered. Without an understanding of the reaction mechanism, investigators have had no way to predict which compounds would be effective in selectively disrupting CCHC zinc fingers.
In view of the foregoing, there exists a need in the art for a mechanism and method which can be used to predict compounds that can effectively disrupt CCHC zinc fingers and, in turn, inactivate the retrovirus of choice.